NMR observability of ATP: preferential depletion of cytosolic ATP during ischemia in perfused rat liver

Murphy, Elizabeth; Gabel, Scott A.; Funk, Alex; London, Robert E.

doi:10.1021/bi00402a003

Cited by 74 publications

(36 citation statements)

References 19 publications

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“…Murphy et al (1988), as mentioned above, have indeed demonstrated that ATP was not 100% NMR-visible in perfused rat liver and have proposed that the invisible ATP pool was mitochondrial. This last proposal was not followed by Hutson et al (1989) who, when working with isolated rat liver mitochondria, reached the conclusion of NMR visibility of ATP and discussed the possibility of the existence of a different intramitochondrial paramagnetic ion environment within either the living organ or the isolated organelles, which could explain the partial observability of ATP in the former situation.…”

Section: Atp Visibility In Perfused C6 Glioma Cellsmentioning

confidence: 80%

“…In this way, both intracellular ADP (Dawson, 1982;Freeman et al, 1983;Iles et al, 1985;Desmoulin et al, 1987) and Pi (Ackerman et al, 1980;Desmoulin et al, 1987) were found to be largely NMR-invisible. On the other hand, ATP was generally been considered to be totally NMR-observable (Cohen, 1983;Stubbs et al, 1984;Iles et al, 1985;Desmoulin et al, 1987;Jeffrey et al, 1989), although the existence of an invisible ATP mitochondria1 pool was reported by Murphy et al (1988). In fact, the existence of different ATP compartments has been clearly demonstrated in adrenomedullary chromaffin cells (Painter et al, 1989), primarily because large variations in ATP longitudinal relaxation time and different pH values for cytosolic and vesicular compartments were found.…”

Section: Phosphorus-31 Nuclear Magnetic Resonance Of C6 Glioma Cells mentioning

confidence: 99%

“…Such an experimental approach avoided any bias linked to the evaluation of the absolute concentration of the nucleotide (particularly with spectra from living cells). Using a similar approach, Murphy et al (1988) demonstrated that in rat liver, ATP was not 100% NMR-visible. These authors analyzed the difference between the relative changes in ATP levels induced by ischemia, when measured either by NMR of the perfused organ or by enzymatic assay with perchloric acid extracts.…”

Section: Atp Visibility In Perfused C6 Glioma Cellsmentioning

confidence: 99%

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Phosphorus‐31 nuclear magnetic resonance of C6 glioma cells and rat astrocytes

Pianet

Merle

Labouesse

et al. 1991

European Journal of Biochemistry

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31P-NMR spectroscopy has been used to study the energy metabolism and the NMR visibility of ATP and intracellular Pi of the C6 glioma cell line and rat astrocyte grown on microcarrier beads with the following results.1. In vivo NMR spectra of C6 glioma cells and rat astrocytes indicate that these cells were able to maintain their level of ATP resonances during a long anoxic period (more than an hour). Both cell types were sensitive to ischemia which induced a loss of ATP resonances within 40 min. Glucose starvation induced a 40% decrease in ATP resonances correlated to a 50% increase in the intensity of the Pi signal. These changes corresponded to a new steady state which could be reversed by reperfusing the cells with a glucose-containing medium.2. In contrast to in vivo data, 31P-NMR analyses of perchloric acid extracts of cells incubated in a glucosefree medium showed that their ATP and Pi contents were unchanged during starvation. The changes of NMR visibility of the metabolites in living C6 cells were correlated to modifications of their macroscopic longitudinal relaxation times, evolving from 0.30 k 0.08 s and 6.6 k 1.5 s in the presence of glucose to 0.68 0.26 s and 3.2 k 0.9 s in the absence of glucose for ATP and Pi, respectively. The changes of the NMR detectability of ATP and Pi indicate that changes in their microenvironment occur during glucose starvation, suggesting the existence of different pools of these metabolites within the cells.3. Under various experimental conditions, i.e. anoxia, ischemia and glucose starvation, rat astrocytes in primary culture showed a very similar behavior to that of C6 cells, suggesting a similar adaptability to the nature of the energy supply for both the normal and the malignant cell. 31P-NMR is expected to be of fundamental interest in cancer research. Numerous studies using this technique have been reported in this field (Daly and Cohen, 1989;Griffiths et al., 1987; Wehrle et al., 1987). In order for NMR to become a useful tool either for improving knowledge of the metabolic processes in cancer tissue or for clinical management, the phosphorylated metabolite status of cells must be analyzed up to the emergence of criteria characteristic of the tumor state. Thus, two basic questions have to be answered regarding the potential usefulness of the technique: (a) do NMR resonances of metabolites in living cells and NMR resonance changes directly reflect their metabolite content and metabolite content changes, respectively; (b) are there significant differences in the energy metabolism between tumor cells and their normal counterparts?As a consequence of the non-invasive character of NMR spectroscopy, the resonance intensities from metabolites within the cells strongly depend on their local environment. In this respect, studies on the NMR visibility of metabolites can provide useful information concerning their intracellular compartmentation. The precise definition of the NMR visibility of a metabolite in living cells implies the determination of the exact amount of met...

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Section: Atp Visibility In Perfused C6 Glioma Cellsmentioning

confidence: 80%

Section: Phosphorus-31 Nuclear Magnetic Resonance Of C6 Glioma Cells mentioning

confidence: 99%

Section: Atp Visibility In Perfused C6 Glioma Cellsmentioning

confidence: 99%

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Phosphorus‐31 nuclear magnetic resonance of C6 glioma cells and rat astrocytes

Pianet

Merle

Labouesse

et al. 1991

European Journal of Biochemistry

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“…In our laboratory, the separation of mitochondria from other organelles in heart tissue was not as successful as has been described for liver [13,161 for which the technique was originally designed. [2,8,101.…”

Section: Discussionmentioning

confidence: 99%

“…Recent studies with 31P NMR have described the preferential depletion of cytosolic ATP during ischaemia in the heart [2] and liver [8]. These conclusions are based on the fact that only the free cytosolic fraction of the total intracellular pool of ATP is NMR-visible.…”

mentioning

confidence: 99%

Subcellular distribution of energy metabolites in the pre‐ischaemic and post‐ischaemic perfused working rat heart

Humphrey

Buckman

Holliss

1990

European Journal of Biochemistry

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Isolated working rat hearts were subjected to 20 min of global ischaemia and either 5 min or 15 min of reperfusion. The subcellular distribution of ATP, ADP, AMP, phosphocreatine and Pi were determined before and after ischaemia by the method of non-aqueous tissue fractionation. Ventricular function and the cytosolic, mitochondrial and ATPase-associated compartmentation of metabolites were measured. After 5 rnin of reperfusion, only 13 k 9% of the pre-ischaemic contractile function was restored compared to 67 f 8% after 15 rnin reperfusion. ATP was reduced in all cellular compartments after 5 rnin of reperfusion but was only decreased from pre-ischaemic values in the cytosolic compartment after 15 rnin of reperfusion (17.1 & 3.9 nmol/mg vs.4.3 k 1.5 nmol/mg total protein; P < 0.05). The mitochondrial [ATP]/[ADP] was reduced from a normal value of 4.36 to 1.79 after 5 rnin but recovered to 4.62 after 15 min of reperfusion. Most of the Pi was located in the mitochondria or with the ATPase fraction of the cell, with only 16% of the total Pi free in the cytosol. This study indicates that the capacity of the heart to recover function may be compromised during early reperfusion by a 59% increase in mitochondrial phosphate content and during late reperfusion by a reduced cytosolic/mitochondrial concentration ratio of both ATP (from 0.85 to 0.19) and phosphocreatine (from 3.9 to 1.24).Myocardial ischaemia is characterised by the depletion of cellular high-energy phosphates, ATP and phosphocreatine. Following the onset of total global ischaemia phosphocreatine declines very rapidly, falling to zero within 5 min as measured by 31P NMR [I, 21, or by 75-85% of the fully oxygenated (normoxic) values as measured chemically [3, 41. ATP, however, declines more slowly and the onset of irreversible damage, in terms of a failure to recover function, is generally accepted to have occurred when ATP has declined to less than 40% of normoxic values [3 -61. After this period, which is between 20 rnin and 30 min in the isolated rat heart [3 -51, the return of flow is associated with a markedly decreased contractile function in spite of the fact that the total tissue content of ATP is restored to approximately 50% of normal and phosphocreatine, often returns to pre-ischaemic values.Compartmentation of reactants can influence the cytosolic phosphorylation potential ([ATP]/[ADP][P,]) which is suggested to be the major control between myocardial respiration and cardiac contractility (for a review, see Gibbs [7]). However, the distribution of ATP and its catabolic products ADP and AMP between the cytosolic, mitochondrial and myofibrillar fractions of the cell following a period of ischaemia is unknown.Recent studies with 31P NMR have described the preferential depletion of cytosolic ATP during ischaemia in the heart [2] and liver [8]. These conclusions are based on the fact that only the free cytosolic fraction of the total intracellular pool of ATP is NMR-visible. However, this is by no means certain and others have reported that all the AT...

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31P Magnetic Resonance Spectroscopy Detects A Functional Abnormality in Liver Metabolism After Acetaminophen Poisoning

et al. 1992

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Eighteen patients with acetaminophen poisoning were studied with 31P magnetic resonance spectroscopy to measure phosphorus-containing metabolites in their livers. The concentrations of all magnetic resonance-detectable metabolites fell in parallel with a decrease in the synthetic ability of the liver, indicated by the prothrombin time ratio (international normalized ratio). In particular, ATP fell to about 20% of its normal concentration in severely affected patients, as did the phosphodiester signal, which is thought to arise mainly from the endoplasmic reticulum in the liver. The correlation between ATP levels and international normalized ratio suggests that the international normalized ratio is a more accurate measure of the number of viable hepatocytes than are other biochemical tests.

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NMR observability of ATP: preferential depletion of cytosolic ATP during ischemia in perfused rat liver

Cited by 74 publications

References 19 publications

Phosphorus‐31 nuclear magnetic resonance of C6 glioma cells and rat astrocytes

Phosphorus‐31 nuclear magnetic resonance of C6 glioma cells and rat astrocytes

Subcellular distribution of energy metabolites in the pre‐ischaemic and post‐ischaemic perfused working rat heart

31P Magnetic Resonance Spectroscopy Detects A Functional Abnormality in Liver Metabolism After Acetaminophen Poisoning

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